The development and homeostasis of the skeletal system depend on the balance between bone formation and resorption. Pathogenic bone resorption modulated by elevation of osteoclasts is a major cause of bone loss in diseases such as osteoporosis, rheumatoid arthritis and osteolytic bone metastasis. Receptor activator of NF?B ligand (RANKL) activates calcium signals that lead to activation of nuclear factor of activated T cells c1 (NFATc1), an essential transcription factor for osteoclastogenesis. Recent discovery of the essential role of costimulatory signals in RANKL-induced osteoclastogenesis provides insights into how RANKL signaling may activate calcium signaling. Costimulatory signals mediated by DNAX-activating protein 12 (DAP12) and Fc receptor common ? subunit (FcR?) cooperate with RANKL to activate phospholipase C ? (PLC?), which leads to changes in intracellular calcium. PLC? has two isoforms, ?1 and ?2. The expression of PLC?1 is ubiquitous, whereas PLC?2 is primarily expressed in hematopoietic cell lineages, among which are the osteoclasts. Consistent with a recent study, we found that the global PLC?2 deficient mice have increased bone mass and decreased osteoclasts. RANKL-induced activation of NFATc1, NF-?B and AP-1 was impaired and in vitro osteoclastogenesis was blocked in the osteoclast precursors derived from PLC?2-/- mice. Further, we found that retrovirus-mediated expression of PLC?2, but not PLC?1, restored RANKL-induced osteoclastogenesis, indicating a unique role of PLC?2 in osteoclastogenesis. However, it is not clear if PLC?2 is required to initiate osteoclastogenesis and/or necessary to maintain osteoclast differentiation. The precise role of PLC?2 in mediating activation of costimulatory signals by RANKL and the necessity of PLC?2 for costimulatory signals to enhance RANKL-induced osteoclastogenesis have not been well characterized. In addition, the global PLC?2 deficient mice have many defects in signaling through the immunoglobulin receptor superfamily receptors, including the FcR? signaling. Thus, osteoclast-specific PLC?2 deficient mice are proposed to be generated in the present studies, so as to characterize a more definitive role of PLC?2 in mediating costimulatory signaling in RANKL-stimulated osteoclastogenesis. Base on our preliminary data and published results, we hypothesize that osteoclast-expressed PLC?2 is required to mediate costimulatory signaling in late stage of RANKL- induced osteoclastogenesis. We will test our hypothesis with three specific aims. Specific aim 1: Define PLC?2-dependent osteoclastogenesis in vitro;Specific aim 2: Characterize the effects of osteoclast-specific PLC?2 ablation on bone phenotype;and Specific aim 3: Determine the role of PLC?2 in RANKL-induced osteoclastogenesis in mice. These studies will define the precise role of PLC?2 in mediating costimulatory signals in RANKL-stimulated osteoclastogenesis. Elucidating the definitive role of PLC?2 in osteoclastogenesis and bone homeostasis will expend our knowledge of costimulatory signaling cascade in osteoclastogenesis and may lead to develop novel strategies and targets to prevent and cure osteolytic-based bone diseases. Pathogenic bone resorption modulated by elevation of osteoclasts is a major cause of bone loss related diseases such as osteoporosis, rheumatoid arthritis and osteolytic bone metastasis. This proposal will generate a tissue- specific mouse model to investigate molecular mechanisms of bone destruction by an important enzyme phospholipase C?2, which will identify novel targets to design drugs to prevent and treat bone loss diseases.